Chromosome 3 comprises approximately 7% of the human genome, or about 200 million base pairs of DNA. Approximately 200 unique sequence probes specific to this chromosome are available, and are being regionally mapped by means of a panel of somatic cell hybrids, each of which contains a rearranged chromosome 3. These two resources permit the construction of a limited map of the chromosome. Two relatively new technologies, pulsed field gel electrophoresis and cloning in artificial yeast chromosomes, will greatly add to the information that can be obtained by conventional means. Pulsed field gel analysis will allow long range restriction mapping around regionally mapped probes; yeast artificial chromosome will allow the isolation, in a single clone, of large stretches of contiguous DNA sequences. The long range goal of this proposal is to contribute to the construction of a complete physical map of chromosome 3 by: 1) constructing a library of large fragments of chromosome 3 in the yeast S. cerevisiae. These fragments will average greater 200 kb in size and will be maintained as artificial yeast chromosomes; 2) screening this library with a minimum of 200 chromosome 3-specific unique sequence probes; and 3) analyzing these clones by pulsed field and conventional gel electrophoresis to determine their sequence content and organization, to develop restriction maps of them, and to obtain probes for the isolation of overlapping clones. It is anticipated that this project will generate much information contributing to the completion of a map of human chromosome 3. This work will be conducted in close collaboration with investigators focussing on the production of additional chromosome 3-containing hybrids, the isolation and characterization of further unique sequence probes, and the detailed analysis of limited regions of the chromosome.